Parasitic nematode (PN) infections remain a major threat to human health worldwide, with more than 1 billion people infected. Children, pregnant women, and the elderly are particularly susceptible to morbidity from nematode infection. Control strategies are restricted to periodic de-worming of infected individuals, which is limited by rapid re-infection rates and the development of drug resistant worm populations. There are no vaccines available for PN infections in humans. Development of new drugs and vaccines will require a better understanding of host factors that participate in the immune response against PN infection. The requirement of a suitable host and the lack of good animal models have limited investigations into mechanisms that animal hosts employ to oppose PN attacks. Here we propose to use a system consisting of three model organisms: an insect, Drosophila melanogaster; the insect and human PN Heterorhabditis gerrardi; and its mutualistic bacteria Photorhabdus asymbiotica. This system is unique because it promises to reveal not only how pathogens evolve virulence but also how two pathogens (worm and bacteria) can synergize to exploit a common host (insect). Despite the identification and characterization of the main NF-?B immune signaling pathways in Drosophila, other evolutionary conserved pathways might regulate host immune mechanisms. It was recently shown that Transforming Growth Factor-beta (TGF-?) superfamily signals modulate the Drosophila immune response to wounding and bacterial infection and we have preliminary evidence suggesting that certain TGF-? pathway signaling molecules are potentially involved in the fly immune response against the nematodes and their associated bacteria. We will use this information to elucidate the exact role of TGF-? signaling components in Drosophila antinematode and antibacterial defense mechanisms. In Aim 1, we will analyze the Drosophila tissuespecific transcriptional regulation of TGF-? signaling members upon infection with Heterorhabditis and Photorhabdus, and the potential interference of TGF-? signaling with other innate immune pathways. In Aim 2, we propose to examine whether TGF-? molecules modulate cellular immune functions and whether the latter interact with humoral reactions in response to the pathogens. In Aim 3, we will explore the involvement of TGF-? signaling in the phenoloxidase/melanization response of Drosophila flies infected with Heterorhabditis axenic or symbiotic nematodes or Photorhabdus bacteria alone. We expect that the results from this project will generate novel insights into the potential role of TGF-? signalng molecules in the host anti-nematode/antibacterial immune response and thus may expose a currently unknown layer of the innate immune system. Such knowledge will contribute significantly to the development of improved practices to control PN in humans.